1. Field of the Invention
This invention relates to a new immunochemical reagent, a new clinical diagnostic test using such reagent, and a test slide using such reagent. More particularly, the invention relates to a new carrier on which globulins useful in clinical diagnostic agglutination reactions may be permanently fixed. The new reagent is particularly useful in agglutination reactions for the detection of rheumatoid factors (RF), and may be advantageously dried onto a test slide to which a test solution may be later applied.
2. Description of the Prior Art
Serologic agglutination reactions have long formed the basis of tests useful in the clinical diagnosis of disease, particularly of bacterial, parasitic, ricksettial and viral diseases. Some of these tests are relatively simple, others require special facilities. Many of the serologic agglutination tests useful today are described in Davidsohn and Wells, Clinical Diagnosis by Laboratory Methods, 13th ed., chaps. 23-24 (Saunders: Philadelphia 1962).
Among the many diseases which may be diagnosed by a serological agglutination test is rheumatoid arthritis. In the serum of the majority of patients having this disease, there exist certain factors--termed rheumatoid factors (RF)--which react in a variety of serologic systems, all of which contain gamma globulin in some form. The exact nature of RF is not certain. Evidence points to their being anti-bodies with specificity for gamma globulin, particularly immunoglobulin G, and certainly they behave as if they were such in serological agglutination reactions.
The agglutinating capacity of RF was first noted in 1940 by Waaler who reported (Acta Path. Microbiol. Scand. 17:172, 1949) on the agglutination of sensitized sheep cells (sheep erthrocytes "coated" with rabbit anti-sheep cell anti-bodies) by rheumatoid arthritis sera. In 1948, this phenomenon was rediscovered by Rose and others and proposed (Proc. Soc. Exp. Biol. Med. 68:1, 1948) as the basis for the diagnostic test known as SCAT--sheep cell agglutination test. This test has since been greatly improved and modified. In 1954, Heller and co-workers demonstrated the reactivity of RF for human gamma globulins when they discovered that Fraction II (FII) human gamma globulin could inhibit the agglutinating property of rheumatoid sera in the SCAT and that tanned sheep cells "coated" with FII human gamma globulin were agglutinated by rheumatoid sera. (J. Immun. 72:66, 1954 and 69:27, 1952). This work was followed by the fashioning of numerous agglutination tests for RF in which red cells or inert particles were coated with antibody or gamma globulin which could then be exposed to and agglutinated by serum containing RF. These tests are referred to at pages 891-894 of Clinical Diagnosis by Laboratory Methods, referred to above, and also in articles by Christian, "Rheumatoid Factors A. Nature and Significance of Rheumatoid Factors" and Cathcart, "Rheumatoid Factors B. Serologic Techniques", both in the treatise Laboratory Diagnostic Procedures in Rheumatic Diseases (Little, Brown and Co., Boston, 1967).
The "perfect" diagnostic test for rheumatoid arthritis has yet to be discovered--one which will be completely specific for that disorder, yet sensitive enough to detect every patient who has definite disease. The most specific serologic tests are those which employ rabbit gamma globulin, such as the SCAT test referred to above. However, these are not as sensitive as tests employing human gamma globulin, such as the FII latex fixation test in which FII human gamma globulin is coated on latex particles. (The latter test is described in the references above and was originally described by Singer and Plotz in the American Journal of Medicine, Vol. 21, p. 888 (1956).) The SCAT has other drawbacks as well. It requires fresh sheep blood cells collected in Alsever's Solution; these cells are stable only for a period of about two weeks and must be washed and brought to a 2%, by volume, suspension for sensitizing with the rabbit anti-sheep red cell antiserum (amboceptor). As the cells age, the amount of amboceptor necessary to achieve a given level of sensitivity may vary. Once sensitized, the cells must be used immediately. These disadvantages have caused the SCAT to become less popular than tests using inert particles instead of red cells on which to coat the antibody or gamma globulin, even though, as mentioned above, the SCAT is the most specific serological test known for rheumatoid arthritis. This lessening in popularity has been aggravated by the fact that it is not possible to use the SCAT in a dry test slide procedure, thus rendering it impractical for rapid screening tests. In the original latex fixation test described by Singer and Plotz the latex and gamma globulin reactants are made only when needed for immediate testing. Finally, a similar problem has been noted in commercial liquid preparations of latex fixations tests, the sensitivity of which shifts over time.
It is an object of this invention to overcome these disadvantages of the SCAT test (and also the disadvantages of the latex fixation tests) by providing a test as specific as the SCAT but having a sensitivity comparable to that of the FII latex fixation test.
It is also an object of this invention to eliminate the necessity of using red cells, which do not have a long shelf life.
It is also an object of this invention to provide a test which may be embodied as a dry test slide for rapid screening and recording of test results.
It is also an object of this invention to provide a test which may be performed easily and which retains its sensitivity over time.